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HIV & HCV
SMART VIRUSES
SMART(ER)- TUBE
A new medical breakthrough for early detection of HIV
& HCV during the window period
Cells– body's basic unit
• An estimated 8.6 million people were living with HIV in Asia in 2006,
including the 960 000 people newly infected in the previous year.
• The highest HIV infection levels in Asia are in south-east Asia, where
combinations of unprotected paid sex and sex between men, along
with unsafe injecting drug use, are fuelling the epidemics.
What is AIDS/ HIV?
• AIDS is a medical condition. People develop AIDS because HIV
has damaged their natural defenses against disease.
• The only reliable way to tell whether someone has HIV is through
blood tests, which can detect the infection -- a few weeks after
the virus first entered the body.
HIV seeks and destroys a type of white blood cell called T cells
also known as T-helper cells or CD4 cells which the body must
have to fight disease.
Without T-helper cells, which kill cells that have been infected with
germs, many other immune system cells including B-cells that
make antibodies, cannot not work properly.
A person infected with HIV may not show any symptoms for years.
But untreated, the number of T-helper cells steadily drops.
Eventually, the numbers fall so low that the risk of infection greatly
increases, and the symptoms of AIDS appear.
How does HIV lead to AIDS
• A damaged immune system is not only more vulnerable to HIV,
but also to the attacks of other infections.
• With time HIV+ve persons may become ill more and more often.
• However, with HIV and HCV, the most devastating and chronic
infection--it could take weeks or up to many months to see any
antibodies in the blood.
• The HIV virus, on the other hand, has its genetic material made
from RNA. It has to insert its genetic code into that of the host
cell in order to replicate. In order to achieve this it must first
make a DNA copy so that it is compatible with the DNA of the
host cell. DNA is then made using the code of the RNA. Since
this is the opposite of the usual case the viruses that do this are
called retroviruses.
What makes the HIV virus different
• HIV is a smart virus that tricks and evades the body's defenses.
Once the HIV virus takes hold, the immune system can never
fully get rid of it.
• HIV+ve persons may look and feel perfectly well for many years
and may not even know that they are infected. But as the
immune system weakens they may become increasingly
vulnerable to even minor illnesses which a normal person could
have been fought off easily.
What is the size of the HIV virus?
An HIV particle is around 100-150 billionths of a meter in diameter.
That's about the same as:
• 0.1 microns
• 4 millionths of an inch
• Replication: Once HIV binds to a cell, it hides HIV DNA inside the
cell's DNA: this turns the cell into a sort of HIV factory.
Retroviruses are not inactive in the latent period
• Retroviruses are unique in biology. Until their classification in the 1970s, it was
believed that all organisms relied on the same fundamental system to turn their
genetic code into proteins necessary for life. The flow of genetic information was
considered to be one way i.e.– code held in DNA was transcribed into RNA and
then translated into proteins.
• This DNA is then spliced into the host cell’s own genes, where they settle down
and wait for the activation of the host cell to make new virus particles.
• After the HIV infection diagnosis is made using a blood test to detect antibodies
to the virus or copies of the virus itself. The length of this period varies from
person to person, and depends on a wide range of factors, such as the amount
of HIV present in the bloodstream, general health, the presence of other
illnesses, and the response to treatment.
• During the asymptomatic period, the virus is far from inactive. It is constantly
replicating and causing damage to the immune system. The immune system is
highly resilient, however, and it takes time for this damage to make an impact.
The
•Whenlifecycle of
HIV virions enter the HIV of a new host, a protein on the surface of the virus
bloodstream
- gp120 develops an affinity for CD4 receptor in the blood called CD4+ or T helper cells.
• When a virion encounters a CD4+ cell there is a reaction, which attaches the virion to the
host cell. This binding is strengthened further by a co-receptor on the cell surface.
• Once the virion has been bound to the host cell surface, its next task is to get inside. This is
achieved through the fusion of the virus coat and the cell membrane. Following fusion, the
genetic material of the virus– i.e. RNA, is released into the cell, along with the viral
enzymes-- reverse transcriptase and integrase.
• Reverse transcriptase reads the viral RNA and builds the corresponding DNA strands. The
DNA copy is known as a provirus. Viral DNA now moves to the cell nucleus, where the
cell’s own genes, also made of DNA, are housed. Another viral enzyme, integrase, splices
the strands of DNA into the host cell genome.
• Secure within the host cell’s genes, the proviral DNA can persist for many years in a latent
state, secretly carrying the genetic instructions for making new virions. When the host cell is
activated – that is it begins to divide – the proviral DNA will be transcribed into RNA, which
is then translated into viral proteins and polyproteins. Together, the RNA and these proteins
then migrate to the inside of the host cell membrane, where they will be assembled into
new virions.
• Among the viral proteins is the enzyme, protease, which cleaves the polyproteins into
smaller, functional proteins, thereby allowing the new viral particles to mature . Following
assembly, the newly formed virions bud from the host cell surface, entering the bloodstream
where they will encounter uninfected CD4+ cells and begin another cycle.
Pathophysiology – how HIV causes disease
• The human immune system is an organ concerned with destroying and eliminating from the
body any organism, substance or particle that threatens the body’s integrity. Like the heart or
the brain, an effective immune system is essential to life. It differs from these organs,
however, in having a wide variety of components dispersed throughout the body and
circulating in the blood.
• To mount an immune response, these various components must communicate and interact in
a precisely orchestrated and organized way. A component with pivotal role in coordinating
this is the CD4+ cell, or T-helper cell. The CD4+ cell is the primary target of HIV.
• The CD4+ cell, also known as the T-helper cell (TH), is a central component of the human
immune response. CD4+ cells bear receptors on their surface, which allow them to pick up
antigens, the specific molecules from the virus or other ‘foreign body’ that alerts the immune
system to the presence of something dangerous. Once the CD4+cells have captured
antigens, they help B cells to make antibodies, and they release lymphokines, chemicals that
stimulate other varieties of immune cell to kill the invader.
• Tc – cytotoxic T cells recognize and destroy cells coated with antigens (the antigen may be
deposited on the surface of a cell as the virus enters, making it a target for Tc activity)
• NK – natural killer cells use the cell-surface changes that result from viral infection to
identify and kill infected cells
• K – killer cells can bind to antibodies, which ‘flag up’ infected cells for killer cells to destroy
• Granulocytes – can engulf and digest infected cells
• Macrophages – release chemicals that stimulate and control the actions of other effector
cells, but also engulf and digest infected cells
Window Period
• HIV and HCV carriers can be identified by detecting the
antibodies against the virus in the patient’s blood. However
antibodies against HIV and HCV take weeks - or months - after
infection to develop in the body. This in essence creates a
‘window period’ in which infected people will be considered non-
infected as long as they do not produce antibodies in their body.
• A test which detects the HIV virus directly is the RNA test. The time
between HIV infection and RNA detection is 9–11 days. These tests,
are more costly and used less often than antibody tests.
Challenges posed by the Window Period
• Medical community has been trying to solve the ‘window period’
problems with modest success. Diagnostic companies are trying
to develop kits to detect lower levels of antibodies and enable
early diagnosis of HIV.
• Some other methods like the p24 antigen test– which look for
proteins or antigens -- have been able to shorten the window
period by a few days but they are not very cost effective.
• A great stride has been in the testing of the viral genome - the
nucleic acid of the virus, which has shortened the window by
about 12 days but it is a very expensive test. For a positive
result there should be virus in the blood which can take weeks
or even months. So, an infected person may test negative.
• Once you are infected, you probably will remain infected for life.
It could take years for you to begin showing the symptoms of
AIDS.
Hepatitis C virus (HCV)
• Is a blood-borne infectious disease caused by the Hepatitis C virus (HCV)
affecting some 150-200 million people worldwide.
• The hepatitis C virus is usually detectable in the blood within one to three
weeks after infection, and antibodies to the virus are generally detectable
within 3 to 12 weeks.
HIV - HCV
• HIV antibody test: shows whether a person is infected with HIV. Antibody tests
are also known as ELISA (Enzyme-Linked Immunosorbent Assay) tests.
• Antigen test.: Antigens found on a foreign body or germ, trigger the production
of antibodies in the body. The antigen on HIV that most commonly provokes an
antibody response is the protein P24. Early in the infection, P24 is produced in
excess and can be detected in the blood serum by a commercial test (although
as HIV becomes fully established in the body it will fade to undetectable levels).
P24 antigen tests are sometimes used to screen donated blood, but an also be
used for testing for HIV in individuals earlier than standard antibody tests. Some
modern HIV tests combine P24 and other antigen tests with standard antibody
identification methods to enable early and accurate HIV detection.
• There are conflicting views about the reliability and soundness of ELISA, for
instance the ELISA test are not specific for HIV. This is why four repeat ELISA tests
plus a Western Blot are required for a diagnosis of HIV in the USA.
• Even the manufacturers of the ELISA test kit have clearly mentioned on the kit that
ELISA should not be used on its own for HIV diagnosis.
• One of the main drawbacks in ELISA test is that p24 proteins cross react with a
wide variety of uninfected human tissue and blood samples from other diseases like
leprosy, malaria and viral infections. Hence the ELISA test may give false positive
results even without the HIV infection.
• The main argument against the Western Blot test -- based on the
antibody reaction mechanism like the ELISA test, is that it is known
to show non-specific positive reaction to a number of diseases like
leprosy or tuberculosis- a parasitic infection and other viral
infections. Hence the Western Blot is not a determinate diagnostic
tool for indication of HIV.
HIV detection tests & their limitations - PCR Test
• The PCR viral load test studies the magnification of tiny HIV viral
particles in the infected blood.
• One of the main drawbacks of the PCR test is that the viral load
can also increase non-specifically due to other viral and
opportunistic bacterial infections as a result the viral load test may
not always be an indicator for the clinical progression of HIV to
AIDS and may lead to incorrect results.
Risks involved with not detecting all the carriers:
S -timulating
M-aximal
A-ntibody
R-esponse
T-ube
SMARTube--The solution
• To enable antibody production, in a small blood sample, within
days from infection, without having to wait for the body to produce
antibodies weeks or months later, & To push for antibody
production at levels that can be detected by current testing.
• The sterile Plasma inside the SMARTube has a shelf life of 6 month when kept at 2-8o c.
• Before use, SMARTube is brought to room temperature and 1ml of whole blood collected in
heparin is introduced into it. This is then incubated for five days at 37oC in a humidified
CO2 incubator. After incubation a sample of the supernatant is removed for testing using
ELISA test or any other detection method.
• To start using SMARTube, even for the first time you don't need any special equipment or
qualifications.
• The test also does not require any specialized training or capacity building. The only
requirement of this test is 1ml of whole blood collected in heparin (transferred in lab, using
sterile pipette etc.). The blood sample inside it then needs to be incubated at 37oC in
humidified CO2 incubator for 5 days. After incubation a sample of the supernatant is
removed for testing (separation occurs by gravity during incubation), using any currently
available method for HIV/HCV testing.
• SMARTube enables antibody production, in a small blood sample, within days from infection,
without having to wait for the body to produce antibodies weeks or months later. This is done by
pushing for antibody production at levels that can be detected by current antibody testing
(ELISA).
• SMART Technology involves in vitro activation of the HIV and/or HCV primed lymphocytes,
pushing them to antibody production even in the phase of active suppression. The result of this
activation is higher levels of antibodies in SMARTube culture using available diagnostic kits.
• The greatest advantage of this cutting edge technology is the flexibility and simplicity of use,
enabling the collection of blood even in remote places. The blood sample thus collected can be
transferred to the SMARTube even a day later, when it reaches the testing lab. Since it is a blood
pre-treatment device, once the blood is treated in the SMARTube it can be tested using any HIV
or HCV antibody ELISA tests. Therefore the labs do not need to change their way of diagnosing
the infection, they only change the way the blood is handled prior to the tests. This makes the
SMARTube very simple to use and with a great return for the money, a safer blood supply and
better detection of infected individuals.
• As a blood pre-treatment device SMARTube also reduces the rate of false positives.
SMARTube - Advantages
•Neeraj Mahajan
Country Head (India)
Eternal Health & Wellness Foundation (USA)
Mob: 9999989066, 9818666863